Biomedical subjects

Max Levin

Publications and source records attributed to Max Levin.

7 recordsLinked to original sources

PubMed2006

Separation of human adipocytes by size: hypertrophic fat cells display distinct gene expression.

Enlarged adipocytes are associated with insulin resistance and are an independent predictor of type 2 diabetes. To understand the molecular link between these diseases and adipocyte hypertrophy, we developed a technique to separate human adipocytes from an adipose tissue sample into populations of small cells (mean 57.6+/-3.54 microm) and large cells (mean 100.1+/-3.94 microm). Microarray analysis of the cell populations separated from adipose tissue from three subjects identified 14 genes, of which five immune-related, with more than fourfold higher expression in large cells than small cells. Two of these genes were serum amyloid A (SAA) and transmembrane 4 L six family member 1 (TM4SF1). Real-time RT-PCR analysis of SAA and TM4SF1 expression in adipocytes from seven subjects revealed 19-fold and 22-fold higher expression in the large cells, respectively, and a correlation between adipocyte size and both SAA and TM4SF1 expression. The results were verified using immunohistochemistry. In comparison with 17 other human tissues and cell types by microarray, large adipocytes displayed by far the highest SAA and TM4SF1 expression. Thus, we have identified genes with markedly higher expression in large, compared with small, human adipocytes. These genes may link hypertrophic obesity to insulin resistance/type 2 diabetes.

Adipocytes↗

PubMed2006

ATP depletion in macrophages in the core of advanced rabbit atherosclerotic plaques in vivo.

The cores of rabbit plaques in vivo are hypoxic, suggesting that ATP depletion due to an insufficient supply of oxygen and nutrients could contribute to macrophage death in atherosclerotic plaques. During hypoxia, however, macrophages maintain ATP levels by anaerobic glycolysis. To directly assess ATP and glucose metabolites in plaques in vivo, we used bioluminescence imaging to map the concentrations of ATP, glucose, glycogen, and lactate in normal and atherosclerotic rabbit aortas in vivo. Hypoxia was assessed with NITP (7-(4'-(2-nitroimidazol-1-yl)-butyl)-theophylline). Normal aortas and plaques <500 microm thick were not hypoxic and had homogenous concentrations of energy metabolites. In plaques >500 microm thick, however, the cores were characterized by ATP depletion, low concentrations of glucose and glycogen, and a high concentration of lactate. A majority of ATP-depleted macrophages within the core were viable but severely hypoxic and glucose depleted. Hyperoxia in vitro reversed the ATP depletion in macrophages in viable areas of the core. Our findings suggest that ATP depletion contributes to the death of macrophages in atherosclerotic lesions and to the formation of a necrotic core.

Adenosine Triphosphate↗

PubMed2005

Adenosine concentration in the porcine coronary artery wall and A2A receptor involvement in hypoxia-induced vasodilatation.

We tested whether hypoxia-induced coronary artery dilatation could be mediated by an increase in adenosine concentration within the coronary artery wall or by an increase in adenosine sensitivity. Porcine left anterior descendent coronary arteries, precontracted with prostaglandin F(2alpha) (10(-5) M), were mounted in a pressure myograph and microdialysis catheters were inserted into the tunica media. Dialysate adenosine concentrations were analysed by HPLC. Glucose, lactate and pyruvate were measured by an automated spectrophotometric kinetic enzymatic analyser. The exchange fraction of [(14)C]adenosine over the microdialysis membrane increased from 0.32 +/- 0.02 to 0.46 +/- 0.02 (n = 4, P < 0.01) during the study period. At baseline, interstitial adenosine was in the region of 10 nM which is significantly less than previously found myocardial concentrations. Hypoxia (P(O(2)) 30 mmHg for 60 min, n = 5) increased coronary diameters by 20.0 +/- 2.6% (versus continuous oxygenation -3.1 +/- 2.4%, n = 6, P < 0.001) but interstitial adenosine concentration fell. Blockade of adenosine deaminase (with erythro-9-(2-hydroxy-3-nonyl-)-adenine, 5 microM), adenosine kinase (with iodotubericidine, 10 microM) and adenosine transport (with n-nitrobenzylthioinosine, 1 microM) increased interstitial adenosine but the increase was unrelated to hypoxia or diameter. A coronary dilatation similar to that during hypoxia could be obtained with 30 microM of adenosine in the organ bath and the resulting interstitial adenosine concentrations (n = 5) were 20 times higher than the adenosine concentration measured during hypoxia. Adenosine concentration-response experiments showed vasodilatation to be more pronounced during hypoxia (n = 9) than during normoxia (n = 9, P < 0.001) and the A(2A) receptor antagonist ZM241385 (20 nM, n = 5), attenuated hypoxia-induced vasodilatation while the selective A(2B) receptor antagonist MRS1754 (20 nM, n = 4), had no effect. The lactate/pyruvate ratio was significantly increased in hypoxic arteries but did not correlate with adenosine concentration. We conclude that hypoxia-induced coronary artery dilatation is not mediated by increased adenosine produced within the artery wall but might be facilitated by increased adenosine sensitivity at the A(2A) receptor level.

Acetamides↗

PubMed2005

Postnatal constriction, ATP depletion, and cell death in the mature and immature ductus arteriosus.

After birth, constriction of the full-term ductus arteriosus induces oxygen, glucose and ATP depletion, cell death, and anatomic remodeling of the ductus wall. The immature ductus frequently fails to develop the same degree of constriction or anatomic remodeling after birth. In addition, the immature ductus loses its ability to respond to vasoconstrictive agents, like oxygen or indomethacin, with increasing postnatal age. We examined the effects of premature delivery and postnatal constriction on the immature baboon ductus arteriosus. By 6 days after birth, surrogate markers of hypoxia (HIF1alpha/VEGF mRNA) and cell death [dUTP nick-end labeling (TUNEL)-staining] increased, while glucose and ATP concentrations (bioluminescence imaging) decreased in the immature ductus. TUNEL-staining was significantly related to the degree of glucose and ATP depletion. Glucose and ATP depletion were directly related to the degree of ductus constriction; while TUNEL-staining was logarithmically related to the degree of ductus constriction. Extensive cell death (>15% TUNEL-positive cells) occurred only when there was no Doppler flow through the ductus lumen. In contrast, HIF1alpha/VEGF expression and ATP concentrations were significantly altered even when the immature ductus remained open after birth. Decreased ATP concentrations produced decreased oxygen-induced contractile responses in the immature ductus. We hypothesize that ATP depletion in the persistently patent immature newborn ductus is insufficient to induce cell death and remodeling but sufficient to decrease its ability to constrict after birth. This may explain its decreasing contractile response to oxygen, indomethacin, and other contractile agents with increasing postnatal age.

Adenosine Triphosphate↗

PubMed2005

ATP depletion and cell death in the neonatal lamb ductus arteriosus.

Postnatal constriction of the full-term ductus arteriosus produces cell death and remodeling of the ductus wall. Using a bioluminescence imaging technique, we found that after birth, the lamb ductus develops ATP, glucose, and glycogen depletion in addition to hypoxia. In vitro studies showed that cell death correlates best with ATP depletion and is most marked when both glucose and oxygen are severely depleted; in addition, the degree of ATP depletion found in vivo is sufficient to account for the extensive degree of cell death that occurs after birth. Under hypoxic conditions, the immature ductus is more capable of preserving its ATP supply than the mature ductus as a result of increased glucose availability, glycogen stores, and glucose utilization. However, the immature ductus is just as susceptible as the mature ductus to ATP depletion when glucose supplies are restricted. The extensive degree of cell death that occurs in the newborn ductus after birth is associated primarily with ATP depletion. The increased glycolytic capacity of the immature ductus may enable it to tolerate episodes of hypoxia and nutrient shortage, making it more resistant to developing postnatal cell death and permanent closure.

Adenosine Triphosphate↗

PubMed2005

Formation of multicellular epithelial structures.

The kidney is primarily comprised of highly polarized epithelial cells. Much has been learned recently about the mechanisms of epithelial polarization. However, in most experimental systems the orientation of this polarity is determined by external cues, such as growth of epithelial cells on a filter support. When Madin-Darby canine kidney (MDCK) cells are grown instead in a three-dimensional (3D) collagen gel, the cells form hollow cysts lined by a monolayer of epithelial cells, with their apical surfaces all facing the central lumen. We have found that expression of a dominant-negative (DN) form of the small GTPase Rac1 causes an inversion of epithelial polarity, such that the apical surface of the cells instead faces the periphery of the cyst. This indicates that the establishment of polarity and the orientation of polarity can be experimentally separated by growing cells in a 3D collagen gel, where there is no filter support to provide an external cue for orientation. DN Rac1 causes a defect in the assembly of laminin into its normal basement membrane network, and addition of a high concentration of exogenous laminin rescues the inversion of polarity caused by DN Rac1.

Animals↗

PubMed2004

Computerized determination of adipocyte size.

OBJECTIVE: Fat cell size is a fundamental parameter in the study of adipose tissue metabolism, because it markedly influences the cellular rates of metabolism. Previous techniques for the sizing of adipocytes are often complicated or time-consuming. The aim of this study was to develop a new, computerized method for rapid and accurate determination of adipocyte size in a cell suspension obtained by incubating human or rat adipose tissue biopsies with collagenase. RESEARCH METHODS AND PROCEDURES: The cell suspension was placed between a siliconized glass slide and a cover slip. Using the reference method [designated as (R)], the cell diameters were determined manually using a microscope with a calibrated ocular. The new method presented here [designated as (C)] was based on computerized image analysis. RESULTS: After two well-defined corrective adjustments, measurements with (R) and (C) agreed very well. The small remaining differences seemed, in fact, to depend on inconsistencies in (R). DISCUSSION: We propose that (C) constitutes a valuable tool to study fat cell size, because this method is fast and allows the assessment of a sufficient number of cells to get reliable data on size distribution. Furthermore, images of cell preparations may be stored for future reference.

Adipocytes↗